DNase IIβ Distribution and Activity in the Mouse Lens

Alicia De Maria, Steven Bassnett
2007 Investigative Ophthalmology and Visual Science  
PURPOSE. To map the cellular and subcellular distribution of DNase II␤ activity in the mouse lens. METHODS. DNase II␤-specific activity was determined by assaying lens lysates prepared from wild-type or DNase II␤-null mice. Regional nuclease activity was determined by microdissection of lens samples or a tissue-imprinting assay. Subcellular distribution was determined by density-gradient ultracentrifugation. RESULTS. DNase II␤ transcripts increased 200-fold in abundance during fiber cell
more » ... g fiber cell formation, and DNase II␤ activity accounted for approximately 50% of the acid nuclease activity in the cortical fiber cells. Examination of lenses from DNase II␤-null mice confirmed that the enzyme was required for denucleation. In wild-type lenses, nuclei were TUNEL positive before denucleation, indicating that 3Ј-OH DNA ends had accumulated. However, DNase II␤-mediated scission generates 3Ј-PO 4 Ϫ DNA ends only. This paradoxical finding was explained by the presence of phosphatases that converted the 3Ј-PO 4 Ϫ ends produced by DNase II␤ into 3Ј-OH ends. DNase II␤ activity was strongest early in differentiation, where it was associated with the lysosomal fraction. Later, an increasing proportion of DNase II␤ activity was found in the cytosol. CONCLUSIONS. DNase II␤ activity correlated with and was necessary for fiber denucleation and was most likely contained initially within fiber cell lysosomes before release into the cytoplasm. (Invest Ophthalmol Vis Sci. 2007;48:5638 -5646)
doi:10.1167/iovs.07-0782 pmid:18055814 fatcat:rgqac6bs25a7ziezl6k7gqa4iu